Project 2: Mechanisms of Androgen Receptor Transactivation in Recurrent Prostate Cancer The androgen receptor (AR) is required for normal prostate growth and development and likely contributes to the development of prostate cancer. Recent evidence suggests that the AR is activated in the absence of testicular androgen in recurrent prostate cancer. Studies using the CWR22 human prostate cancer xenograft indicate that despite low levels of circulating androgen in the castrated male, AR can be reactivated through growth factor-induced signal transduction. The goal of this research is to understand the molecular mechanisms involved in AR mediated growth of recurrent prostate cancer. Reactivation of AR in the presence of low androgen levels appears to be driven by mitogen signaling. We propose to further test this hypothesis through studies on the molecular mechanisms involved in epidermal growth factor (EGF) or transforming growth factor a (TGFa) and heregulin (HRG) stimulation of AR transactivation in recurrent prostate cancer ceils. Our ongoing studies provide strong preliminary evidence that mitogen-induced receptor tyrosine kinase signaling increases AR transactivation by direct effects on AR and/or AR coregulators that are only partially blocked by AR antagonists. In Aim 1 we will investigate mechanisms by which EGF/TGFa and HRG increase AR transcriptional activity in recurrent prostate cancer. We will determine the effects of these mitogens on AR phosphorylation, the role of p160 coactivators in mitogeninduced increases of AR transactivation, and the composition of the EGF/TGFa and HRG-induced AR transcription complex. In Aim 2 we will establish the role of mitogen signaling in the increased sensitivity of recurrent prostate cancer cells to androgen-induced proliferation. We will identify downstream kinase signaling mechanisms of EGF/TGFa and HRG involved in AR activation. In Aim 3 we will determine the AR domains involved in EGF and HRG-induced interactions with AR coactivators and corepressors and identify mechanisms by which these mitogens interfere with transcriptional repression by AR antagonists in prostate cancer. We will focus on determining the role of AR activation function 3 and coactivators and corepressors in mitogen enhanced AR transactivation to establish the molecular basis for loss of inhibition by AR antagonists. In Aim 4 we will identify other coregulators that interact with AR in response to EGF/TGFa and HRG signaling. We will determine the effects of these mitogens on the interaction of coregulators with the AR NH2-terminal or DNA binding domain and identify novel proteins that interact with AR in response to EGF signaling. The proposed studies will improve our understanding of molecular aspects of AR activity in prostate cancer by revealing mechanisms that contribute to the recurrent growth of prostate cancer in the androgen deprived patient.